Heteroduplex mobility assay (HMA) pre-screening: An improved strategy for the rapid identification of inserts selected from phage-displayed peptide libraries
Fred Fack
Laboratoire National de Santi, P.O. Box 1102, 20A, Rue Auguste
Lumihre, L-1101 Luxembourg, Luxembourg
Sabrina Deroo
Laboratoire National de Santi, P.O. Box 1102, 20A, Rue Auguste
Lumihre, L-1101 Luxembourg, Luxembourg
Stephanie Kreis
Laboratoire National de Santi, P.O. Box 1102, 20A, Rue Auguste
Lumihre, L-1101 Luxembourg, Luxembourg
(Author for correspondence e-mail:
kreis@cu.lu)
Claude P. Muller
Laboratoire National de Santi, P.O. Box 1102, 20A, Rue Auguste
Lumihre, L-1101 Luxembourg, Luxembourg
Abstract
Phage-displayed peptide libraries represent an efficient tool to
isolate peptides that bind a given target molecule. After several
selection rounds, generally a large pool of target binding phages is
obtained. Conventional analysis of the selected phage population
involves extensive sequencing of many clones, most of which can be
identical. We have adapted the Heteroduplex Mobility Assay (HMA)
for pre-screening of phage inserts that were amplified by direct
colony PCR of ELISA-positive clones. This strategy allowed for the
rapid and reproducible assignment of insert sequences to different
'heteroduplex migration groups'. Sequence analysis of only one
representative of each HMA migration group then completes the
characterisation of the binding phage population. In our model
experiments, only 16% of HMA pre-screened clones required
further sequence analysis.
Keywords
DNA conformation, heteroduplex mobility assay (HMA), mismatch,
mutation detection, phage display
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